APS - Tecnologia Genetica -

Prévia do material em texto

Trabalho de Tecnologia Genética: Diagnostico Molecular e Bioinformática = APS
Aluno: Clebson Barbosa carvalho RA: 20623517
Professor: Daniele Gonçalves Castilho 
struções para a APS: 
A técnica de PCR (Polymerase Chain Reaction – Reação em Cadeia da Polimerase) é atualmente amplamente utilizada para o sequenciamento de genes e diagnóstico de doenças hereditárias, identificação de impressões digitais do material genético e detecção e diagnóstico de doenças infecciosas. Por ser um método direto de alta sensibilidade, a reação em cadeia de polimerase está sendo considerada uma grande aliada na detecção de doenças infecciosas. Sua versatilidade, que permite a aplicação em diversos materiais biológicos, aliada à rapidez de análise é outro diferencial da técnica (http://www.cjflash.com.br/cace/informe,biologia-molecular-porpcr,7.html).
ATIVIDADE 1:
1) Assista os vídeos disponíveis nos links:
PCR: https://pt.khanacademy.org/science/biology/biotech-dna-technology/dna-sequencing-pcrelectrophoresis/v/the-polymerase-chain-reaction-pcr
Eletroforese: https://pt.khanacademy.org/science/biology/biotech-dna-technology/dna-sequencing-pcrelectrophoresis/v/gel-electrophoresis-dna
ATIVIDADE 2: Leia os textos disponíveis nos links:
Reação em cadeia da polimerase: https://pt.khanacademy.org/science/biology/biotech-dna-technology/dnasequencing-pcr- electrophoresis/a/polymerase-chain-reaction-pcr
Eletroforese em gel: https://pt.khanacademy.org/science/biology/biotech-dna-technology/dna-sequencing-pcrelectrophoresis/a/gel-electrophoresis
Sequenciamento de DNA: https://pt.khanacademy.org/science/biology/biotech-dna-technology/dna-sequencingpcr- electrophoresis/a/dna-sequencing
ATIVIDADE 3: O aluno deverá copiar as questões abaixo, colar num documento de preferência, e responder qual a reposta correta.
1- Qual seria o fluxo em um laboratório de biologia molecular para se obter o resultado de um paciente que está realizando uma pesquisa de bactéria por PCR?
X a) Extração de DNA --> PCR --> Eletroforese
b) Eletroforese --> PCR --> Extração de DNA
c) PCR --> Eletroforese --> Extração de DNA
d) Eletroforese --> Extração de DNA --> PCR
e) PCR --> Extração de DNA --> Eletroforese
Resposta: a)
2- A reação da PCR consiste em amplificar um fragmento (região específica) de DNA milhões de vezes, para que isso ocorra precisamos preparar a reação previamente. Que reagentes são necessários para a montagem de uma reação de PCR?
X a) água, buffer, MgCl, DNTPs, Taq polimerase, primers e DNA
b) água, buffer, helicase, DNTPs, Taq polimerase, primers e DNA
c) água, buffer, Mgcl, DNTPs, DNA polimerase, primers e DNA
d) água, buffer, helicase, DNTPs, DNA polimerase, primers
e DNA e) água, buffer, primase, DNTPs, Taq polimerase, primers e DNA
Resposta: a)
3- Qual é o propósito para o qual a PCR é usado?
a) Analisar a expressão gênica do DNA
b) Extrair o DNA de células
X c) Amplificar em fragmento específico de DNA
d) Fazer todo o processo de replicação igual a célula
e) Analisar o tRNA presentes nas células
Resposta: c)
4- O que acontece com o DNA quando o termociclador atinge a temperatura de 90-95 ° C por 30 segundos no primeiro passo da PCR?
a) Os primers se ligam na fica única de DNA
b) A taq polimerase sintetiza as novas fitas do DNA, utilizando os nucleotídeos provenientes da DNTP
c) Ocorre a desnaturação de proteínas
X d) As fitas de DNA se separam
e) Ocorre a renaturação da molécula de DNA
Resposta: d)
5- As seguintes etapas constituem a reação em cadeia da polimerase (PCR), onde milhões de cópias do fragmento de DNA que se quer amplificar são gerados,
I. Desnaturação do DNA genômico (94ºC, 30s).
II. Anelamento dos iniciadores no DNA genômico (55-65ºC, 30s).
III. Desnaturação do DNA genômico (94ºC, 5min).
IV. Síntese do DNA pela polimerase (72ºC, 2-5 min).
A sequência correta da reação apresentada está na opção:
X a) I - II - III - IV
b) III - II - IV- I
c) II - I - IV- III
d) IV - I -III-II
e) III - I - II - IV
Resposta: a)
Desenhar Primers:
SAP-PADG_0
0634
50D-PADG_0
7418
FORMAMIDASE-
PADG_00315
Hypothetical
Protein-PADG_
04791
Glutamate cysteineligase-
NM_001203879.2
Seqüência
Temp. Melting 
62
62
62
%GC
 
 50
 
 50
47,6
Tamanho do primer
 20
 
 22
21
Comprimento
aplication
312
 
 804
	
Seqüência 
SAP-PADG_0
0634
50D-PADG_0
7418
FORMAMIDASE-
PADG_00315
ATGGCAACCCAAATCCAACAAAAGCTCCAAGACAGAAATGCCGGATATGCAAACACTTTCAATCAGAGTCATCTTGCCATTCCCCCTGTGGAGAAATATCTTGTTCGTATGTTAACCCATACCCCCTCTCCATTGCCCCTCTAGTAACTCTTCAACAGTGACCTGTATGGATGCCCGTATCGACCCCTCCTCCGCCTTTGGCATCTCCCTTGGAGATGCCCACGTAATCCGTAATGCCGGCGCCAGCGCCCGCGATGGCTTGCGCTCCATTGTCATCTCGCAGCAGCTGCTAGGAACCCGCGAGATCCTGCTCATCAAGCACACGGGCTGTGGCATGCTGACCTTCACCAACGAGCAGGCGCATGCTCTTGTTGAGAAGAAATTGGTGGAGAGTCACAGGCAGAATGGAACTGGCGCTGCGGATGGCGGGTGTAGGGTGCTCGAGGCGCTGAAGAGGGATATTCCGGATTTCCAGCCGTTTCCACTGCTT
GAGGAAGCGGTTAGAGAGGATGTTAGGTGGCTGAGTGAGCAGGTGCTGGTTACGCCTGGGATTCCAATTTCCGGGTGGGTGTATGAGATTGAGACGGGGAAGGTGAGGAAAGTTGTGTAG
Hypothetical
Protein-PADG_
04791
ATGTACATGATCATACGAAGAAACACGTTCACTGGTGTCCATGATCAAAATTTAATGTTTTACGTAAAAA
GTACAGATGTTAACTGTTTAGTTTAA
Glutamate cysteineligase-
NM_001203879.2
ATTTTCGTTCATCAGAACCAAGATTTTTTTTGTCGACACAATCAAACCAAATAGATAATGATACTGTTTTTTATAGTAGTATATATATATTTCTTTTCCTATAGAGTATTTTATCTGAGGTCATGTGGTTGGTTAGTGACACACTTTTCATTATTTTACGTTCTAAATTAAATATCGTAAAGTTAAACCTCGTTTAAAGTATACTATAAAGAATAGAAATATCATTAATTCATTATACACACTATAAAGTTAACCTTTGTTTAAACTACTAATAAACTGGTTATTTAAAAAACTATTAGGGTTAAATTTGTGAGAAATGGAAAAGTAAGAGGCAAAAAACGAAAGAATCTTTT
ATTACAATGATTATTAATTTTATCAACTTTCTTTTATAGACAGCTCAGCTCCACTGATCGGCTCATGCTCCACTGATCAACTTTCTTTCATTTTTTTATCAGTTTCTCATTTCTCTCTCAGTCGCATATAAATTTGAAAC
GCCGATTCGTTTTTTGCTCTTTAAGCTTTCTTCTTGATTTCGACGGTGTCTCTCAATCTCCGTCAAGCTTG
ACGAATTTCAGGTGTGATTTGCTTCTCTTCTTCTTCGCTTTCTGGACTTGTGGATTTTGGTTTTTTGATTG
ATTTGTTCTGCTTTTTTTTTTTTGGTTGAATCTTCGCTTTTTCTCATGATTTTTTTTTTGGCTGTTGCATG
ATTTTGGTTTTTGTTAGAGTCGAATTTTTTGCAGATTTTTGCTTTGATTTGATTTTTGTTGTTGTTGTTGTT
TCTGGTTTTGGTGATACACGAATTTTGATTTGTTCTTCCAGATGATAGTTCAATTCTGTGGTAGCTGCTGA
TTCTAATTATGTGATATATGTTATTAGAATTGAGAAAAATGCAGTTATTTGTGATTTCTCATAGCTTATG
ATGAATTGTTATAATTGCAGGAGCTATATATACCATGGCGCTCTTGTCTCAAGCAGGAGGATCATACACT
GTTGTTCCTTCTGGAGTTTGTTCAAAGGCTGGAACTAAAGCTGTTGTTTCGGGTGGCGTGAGGAATTTGG
ATGTTTTGAGGATGAAAGAAGCTTTTTGGTAGCTCCTACTCTAGGAGTCTATCTACCAAATCAATGCTTCT
CCATTCTGTTAAGAGGAGTAAGAGAGGGCATCAATTGATTGTTGCGGCAAGTCCTCCAACGGAAGAGGCT
GTAGTTGCAACTGAGCCGTTGACGAGAGAGGATCTCATTGCCTATCTTGCCTCTGGATGCAAAACAAAGG
ACAAATATAGGTACAGTTTGATGCTTATATGTTCTGTTAAAATGCTACTTAGTGTTTGTAATTGTGACAT
ATTTCTTTGTTTGTGCTCTTTTACGGTGTTGGTATTGTTTGAATTCATCCAGATTGTTTGTCTATTGGAG
AAATATTCTGTTTTTGCTAACTTAACATTGTGAGTTTTCAGAATAGGTACAGAACATGAGAAATTTGGTTT
TGAGGTCAATACTTTGCGCCCTATGAAGTATGATCAAATAGCCGAGCTTCTTAATGGTATCGCTGAAAGA
TTTGAATGGGAAAAAGTAATGGAAGGTGACAAGATCATTGGTCTGAAGCAGGTTAACATTCTCCTGTGTT
TCATCTTTTGAGCCGTTTATTATTTATATACATTTTCAGCCTGTAGAACATGTTAGCTTCATATACAACA
CAATGATCGGTATATTATAAGGTTTCAGTCTTTTTTTTTTTTTTTGCTGTAGATAACTCGTTCTGTCCTTT
TGTTGACCAGGGAAAGCAAAGCATTTCACTTGAACCTGGGGGTCAGTTCGAGCTTAGTGGTGCACCTCTT
GAGACTTTGCATCAAACTTGTGCTGAAGTCAATTCACATCTTTATCAGGTAATTTCAAAGATAATTATTG
CCAATGCTCTCACCCTAAAACGTGACAAATAACCAACTATCTACGGATGTCACAGGTAAAAGCAGTTGCT
GAGGAAATGGGAATTGGTTTCTTAGGAATTGGCTTCCAGCCCAAATGGCGTCGGGAGGATATACCCATCA
TGCCAAAGGTATGACGCTTTCTCTATTCTTAAGATTGAAGAAAGTTAACATCTGTTTGTTTCTAATTCAG
ATAATATTATCTGATTCTACATAGCCAACGTTCGGATTATTTCTTGGTGTTTGTGTTAAGTTTTCCTATG
ACGAGTCAGGCAGTAAGATTTGACATAGTGTAATGTGTTCTGTGCAGGGGAGATACGACATTATGAGAAA
CTACATGCCGAAAGTTGGTACCCTTGGTCTTGATATGATGCTCCGAACGTGTACTGTTCAGGTTAGATTA
ATATTCTGACTTTGCGCCTTTTCATAATTTCCTTAAACTAAACTTTCCAGTTCTCATCAATAATCTTATT
TTGAAACTTATAGAGAAACTGCATTACTTGCAATCTTGACAGGTTAATCTGGATTTTAGCTCAGAAGCTGAT
ATGATCAGGAAGTTTCGTGCTGGTCTTGCTTTACAACCTGTGAGAAGCTCGATCTCTTTGTGGCATAACC
AGAAACTTTTATTAACGATTTTGACTAACCATTGCCTCACTATTATCTAAATTCAACAGATAGCAACGGC
TCTATTTGCGAATTCCCCTTTTACAGAAGGAAAGCCAAACGGATTTCTCAGCATGAGAAGGTATCTAGAT
TCAGTTGTTATGATCCCCGATATCTTTTTGAATTAGTTTCCCTCCCACCTTATAACTCTGGTATGTTTTG
ACAGCCACATATGGACAGACACTGACAAGGACCGCACAGGAATGCTACCATTTGTTTTCGATGACTCTTT
TGGGTGAGTATCGATGTTTAATCTATGAAAAACAATTACATTACATTGGTTGGTTTGGTATGAGAAGCTTCTCAAAAACCAATGACTCTGTCTCGCATTCTTGATGAGTTATTGGAAAATTGATATGTGCTATTCTGTGG
ATTTTCTTACATATGTTTGTGTTCAAGATAGAAAGATGCTTTGGATCTTTATCCTTCTCTTGAAACAGCA
TGGCTAATCTTAATTTTATTGCTCTCTTTTGTTCTTATCTGGTATAGGTTTGAGCAGTATGTTGACTACG
CACTCGATGTCCCTATGTACTTTGCCTACAGAAAGAACAAATACATCGACTGTACTGGAATGACATTTCG
GGTCAAAATCTATCCTTAATCTTTTCTGTTAGGTTCTATAACTGTTTCATTGCAAGGAATTGGCAAATTG
AAAACCATGGGATCTTCTTGAATTATATCTAATCTCATAATATTCTTCATGCTGCAGCAATTCTTGGCTG
GAAAACTTCCCTGTCTCCCTGGTGAACTGCCTTCATATAATGATTGGGAAAACCATCTGACAACAATATT
CCCAGAGGTTAATCATCTGCCTCAAGAGTCTTTCAATCAGCCTATGACTTTCATTTCCCTGTCTTATGTT
GAAGTCTGACAACGTTCCATAACACTAGGTTCGGTTGAAGAGATACTTGGAGATGAGAGGTGCTGATGGA
GGTCCCTGGAGGAGGCTGTGTGCCCTGCCAGCTTTCTGGGTAACTACATCTATTAGCACATATATATTTC
GTGCTTAATGAGAATGTTTCAAAAAGTATTGACTGTTGTTTTGATTTTATCTACAAATTTCAGGTGGGTT
TATTATATGATGATGATAGTCTCCAAGCTATCCTGGATCTGACAGCTGACTGGACTCCAGCAGAGAGAGA
GATGCTAAGGAACAAAGTAATAACAAGACGCACCACATTTACATTTTCTTATTCCACAGTTAGAAAACAT
CTTGTAATAAACTAATACTTGATTTCCAACAGGTCCCAGTTACTGGCTTAAAGACTCCTTTTAGGGATGG
TTTGTTAAAGCATGTCGCTGAAGATGTCCTGAAACTCGCAAAGGTGTGTTTGTATGAACCCTCTTAGAAA
CTTCGAGCCAGCTTTCTATCCCGTCTGGTCCAGCTTCTAACTATTGGACTTATTTTCTCTTTGTTTCTCA
GGATGGTTTAGAGCGCAGAGGCTACAAGGAAGCCGGTTTCTTGAACGCAGTCGATGAAGTGGTCAGAACA
GGTAAATGAAGTTTCCTAATGCATAATTGAATTCCACACCGACATTTTGTTTGATAACCTGGTGGTGAAA
TTTGGTTTTTGAAATGACAGGAGTTACGCCTGCGGAGAAGCTCTTGGAGATGTACAATGGAGAATGGGGAC
AAAGCGTAGATCCCGTGTTCGAAGAGCTGCTGTACTAAGAAAATGGGACGTGAACAAAAGGTGTCTATAA
ACCTTTGGGTGTGAGTTTATGCTATCTGAAGAATTCGAGTCTCCGGAATAAGGATTTTTTTTTTTTTGTTGT
AATCGGATTTTAAAAACTGATTTTGTTTTAGAAATTCGAAGCATTGAAAAGCAGAAGAAAAATTGTATGT
ACTAAACGATTTCGGTGTGGGAAATCGTTTGGGAGGGTGTGTTTGGATCTTGAATAAATTACCCATTTTT
CTTGTCACAAATTTTGTCTACATTTAACGAAATAACTCAAAACTGATTTCAAGGCAAAGGCGACGGATGGG
ATAATCTAGGCTGTGTAGTGGAAAGTTTTTGATGTTTAGTAATTTGGCAATGTATGTTTTTTTTCTAAATGG
GAGAGCATCTTCGATGTAGACATTTACGAGCTTTTCCTTTAATAGGTTTTTGGCTAGTGAAGAACACTGCT
TTCATAATTGAATCCACCAGACTCCGCGTAAACTAAACTGATTACAACACGATGATCCTGGCCGTCGGTA
ATTATCGTAAAACAAGAAGATAGTGGCACTTAGGTGACAGGATAATATGTAAACCCTACCAAGGCGTCCC
TACAAAATGATGAGTGATGACTGATGAGAGTGAGAAAAACCGAGTCTCCACAGTCCACACACTGCCTTAC
CTAATTAATATACCCACCTTTCATTCATGTATGTTACGTCAACGTTTTTATGTCCGTTAAAACTGTAATC
TACATAAGAGATCATAATTTTTAAAATCTCCATGTATTTCGTTGATACTTTAATTTGGCACGTTTCTAGT
TTTTGAGATCAAATGTTTTTAATCTGCTTCTAGTCTTATTTTTCCTAAACCTTAAACTGTAAACCAATTG
CTTTTTCACATAACCTTGCTTGTTTTCAATTTTAAAATTTAAGTCCAATATTTGAAACAACCTAACAAAT
GTCGTCTATGGCAACCAAAAAAAAATAAAAAATCAAAACAATTTGCTAAAAAGGAAATTGATTAAAATTG
ATTTCTTTCATTTAAATTGGAATTTAATATATTAGGTAAACATGTAAACGCAATCGCAATTTTTATGAGA
GACTTCGGGCCGGTTCGTTTAAGTGTCGCTGCGACTATAACTAGCGACTGGCTTTAAAAACGACTGAGTT
TTGTTTGTTTTTTATTGTTTTTTTTTTAAGTTCGAATATGTCGTATAGATCTGTAAAATAATTCAAATGTG
GCTTTGTCCCATAGATTGGTTTTGATTATGCAATCATCTATCACAACAGTGGAACATTATGCAATCATTT
ATTGTCTATCAAATTAATTATGGTTATGA